Ameliorative Potential of Methanol Extract of Tithonia diversifolia (HEMSL.) Leaf on Lead Acetate-Induced Reproductive Toxicity in Male Wistar Rats

Abstract

Prolonged exposure to lead induces oxidative stress which is implicated in male infertility. Tithonia diversifolia Leaf (TDL) has been reported in folkloric medicine to possess anti inflammatory properties. There is dearth of information on the effects of TDL on lead induced male reproductive dysfunctions. Therefore, the effects of Methanol Extract of TDL (METDL) on Lead Acetate (PbA)-induced reproductive toxicity were investigated in male Wistar rats.` Tithonia diversifolia leaves were collected from Botanical Garden, University of Ibadan, and identified at Forestry Research Institute of Nigeria, Ibadan (FHI No.: 110652). The leaves were air-dried, pulverised, macerated in methanol, and the filtrate was concentrated using rotary evaporator. Identification of the phytochemical constituents present in METDL was done by Gas Chromatography Mass Spectrometry (GC-MS). Forty male Wistar rats (120-150 g) were assigned into eight groups (n=5) and treated thus: Group 1 (Distilled Water, 5 mL/kg), Group 2 (15 mg/kg PbA), Group 3 (50 mg/kg METDL), Group 4 (15 mg/kg PbA + 50 mg/kg METDL), Group 5 (100 mg/kg METDL), Group 6 (15 mg/kg PbA + 100 mg/kg METDL), Group 7 (50 mg/kg phytol) and Group 8 (15 mg/kg PbA + 50 mg/kg phytol). Twenty-four hours after the last administration, blood samples were collected via cardiac puncture from the animals under anaesthesia (50 mg/kg sodium thiopental, i.p.). Serum Gonadotropin-Releasing Hormone (GnRH), Follicle Stimulating Hormone (FSH), Luteinizing Hormone (LH), testosterone, nitrotyrosine, Interleukin-1β (IL-1β) and Tumour Necrosis Factor Alpha (TNF-α) were measured using ELISA technique. Testis and hypothalamus were harvested for spectrophotometric analysis of Malondialdehyde (MDA), Superoxide Dismutase (SOD), catalase and reduced glutathione. Histological analysis of parafine-embedded testis and hypothalamus was done via Hemotoxylin and eosin staining technique. Semen analysis was done using Computer Aided Sperm Analyser (CASA). Data were analysed using Students‘ t-test and ANOVA at α0.05. Phytol (3,7,11,15-tetrametylhexadec-2-en-1-ol) had the highest percentage (14.7%) of all the compounds that have antioxidant properties identified in METDL. Sperm motility, livability and counts were significantly increased in Groups 4 (78.8±1.8%; 66.0±3.3%; 383.2±2.1x106 cell/mL) and 8 (75.0±3.2%; 66.0±4.0%; 78.0±3.7 x106 cell/mL) compared to Group 2 (46.4±1.1%; 42.0±3.7%; 34.8±1.3 x106 cell/mL). Testicular SOD (31.2±1.5 vs. 17.3±0.9 Unit/mg tissue) and catalase (2.5±0.2 vs. 0.9±0.2 Unit/mg tissue) were significantly increased in Group 4 compared to Group 2. Testicular and hypothalamic MDA were significantly decreased in Group 8 (0.4±0.0 µM/mg protein; 0.2±0.0 µM/mg protein) compared to Group 2 (0.5±0.0 µM/mg protein; 0.5±0.0 µM/mg protein). Serum TNF-α, IL-1β and nitrotyrosine levels were significantly decreased in Group 8 (63.0±2.0 pg/mL; 433.8±1.8 pg/mL; 3.5±0.2 ng/mL) compared to Group 2 (95.5±1.9 pg/mL; 683.8±2.3 pg/mL; 7.5±0.2 ng/mL). Significant increase was observed in testosterone, LH, FSH and GnRH in Groups 4 (0.2±3.2x103 ng/mL; 4.2±1.2 mIU/mL; 0.7±0.1 mIU/mL; 1260.0±5.4 pg/mL) and 8 (0.1±3.5x103 ng/mL; 5.1±1.1 mIU/mL; 0.8±0.1 mIU/mL; 1319.0±3.6 pg/mL) compared to Group 2 (0.1±3.8x103 ng/mL; 1.7±0.7 mIU/mL; 0.4±0.0 mIU/mL; 1056.0±3.9 pg/mL), respectively. Histology of the testes and epididymides showed cyto-architectural distortion only in Group 2. iii Tithonia diversifolia alleviated lead acetate-induced reproductive toxicity via the mitigation of oxidative stress. The methanol extract also prevented inflammatory response to lead acetate toxicity.